Rapamycin treatment is the first drug intervention to reliably increase mammalian lifespan by 10% or more. Excitingly, this treatment was effective in heterogeneous mice when initiated at 20 months of age, roughly analogous to human beings at 60 years of age. The 3 independent aims of the current project identify genes regulating health benefits of rapamycin treatment, as well as key genes controlling normal mouse aging. This project is made possible by a revolutionary new mouse model, the diversity outcross (DO), developed by Gary Churchill (Co-Investigator). Compared to other models, the genetic diversity of the DO is more than 4 fold greater, as it was derived from 8 highly diverse inbred strains, including 3 wild-derived M. musculus subspecies. Because the DO is an advanced intercross, it has a high density of recombinations that makes candidate gene identification >10 fold more precise than with a standard F2 or N2 cross. We will initiate treatment in 20-month-old DO mice, because old individuals may respond to treatments differently than young, and to model humans who often would not start treatment until about 60 years of age. Sufficient marker alleles will be tested in each individual mouse to map loci associated with physiological aging (immune, renal, cardiac and metabolic) and lifespan. Dense mapping will identify candidate genes. Aim 1 tests the hypothesis that rapamycin benefits mammalian healthspan (healthy lifespan) via the mTOR pathway. This hypothesis predicts that aging in rapamycin-fed DO mice will be influenced by alleles of mTOR pathway genes. If this hypothesis is verified, we will identify the genes. If this hypothesis is rejected, we will identify alternative loci containing genes, such as drug metabolism genes, that govern rapamycin benefits. Aim 2 tests the hypothesis that the mTOR pathway governs normal aging. This hypothesis predicts that the same genes regulating aging in rapamycin-treated DO mice (Aim 1) also regulate aging in littermate controls. If this hypothesis is verified, we will identify genes with natural variants in the mTOR pathway that provide potential targets for clinical treatments with minimal adverse side effects. If this hypothesis is rejected, we will identify alternative loci containing genes that regulate aging, which would suggest novel mechanisms and novel potential clinical treatments. If, in Aim 1, we do not find genes related to benefits of rapamycin, we will combine Aim 1 and Aim 2 data to confirm and extend detection of key genes regulating normal aging rates. Aim 3 tests the hypothesis that immune, renal, cardiac and metabolic aging, as well as healthspan, are regulated by the same loci. If this hypothesis is verified, we will identify genes that have pleiotropic effects on aging in diverse systems. If this hypothesis is rejected, we will identify genes that regulate aging in each individual system;such specificity in genetic regulation of markers of inflammation could provide the means to decouple benefits of rapamycin treatment from its immune suppressive effects. Whether or not the hypothesis is verified, gene identification will suggest clinical treatments. ) PUBLIC HEALTH RELEVANCE: In the US, two of the most critical medical issues we face are the emotional and financial costs related to ill health in the elderly, which is increasing as the "baby boomer" population ages. In addition to the humanitarian benefits of postponing disease, increases in healthy lifespan will have enormous financial benefits for the US. Dietary rapamycin in mice, started at 20 months of age, increased their healthy lifespan by 15% - equivalent to increasing life expectancy of 60- year-old humans by 10 years. The current study will identify genes whose alleles regulate effects of rapamycin and healthy aging in general. This information will target pathways for interventions to retard aging and extend healthy lifespan in human beings.